This application proposes the functional characterization of mycobacterial sigma factors which the PI believes will be vital in facilitating regulated, rapid responses to the changing environments experienced by the bacilli during the course of their life cycle. The application is a continuation of an R29 award during which the PI identified sigma F (sigF) as an important regulator of loci implicated in stress and infection maintenance. Scanning of the MycD database reveals 13 putative sigma factors of which 10 are of the extracytoplasmic function group which in other systems have been shown to regulate cell wall synthesis pathways and protein secretion. The PI proposes to extend his studies by knocking out all of these sigma factors in both MTB and MAC. The knockouts will be characterized for both in vitro and in vivo phenotypes and the identity of sigma factor-dependent genes will be determined by microarray studies conducted on the knockout strains.